The position of the nucleus is carefully controlled in a wide variety of cell types. Nuclear migration plays a role in normal cell migration events and metastasis;defects in nuclear migration lead to the neurological disease Lissencephaly. Nuclear anchorage functions in the development of the neuro-muscular junction and may contribute to muscular dystrophy. A group of three conserved nuclear envelope proteins are required for proper nuclear positioning in C. elegans. Our objective is to characterize how these proteins function to control nuclear positioning and to identify other proteins that function with or in parallel to them. Our central hypothesis is that UNC-84 functions at the inner nuclear membrane to recruit UNC-83 and ANC-1 to the outer nuclear membrane. Together, they bridge the nuclear envelope to connect the nuclear matrix to the cytoskeleton. Our first aim will determine the topology of these three proteins using an in vivo protease protection assay and immuno-EM. Aim 2 will use molecular genetic techniques to test the central link of our model, the interaction between the SUN domain of UNC-84 and the KASH domains of UNC-83 and ANC-1. In aim 3 we expect to link UNC-83 to the cytoskeleton by identifying interacting partners through biochemical and molecular screens using essential portions of the novel domain of UNC-83. In aim 4 we take a genetic approach to identify additional proteins involved in nuclear positioning by cloning existing enhancer of unc-83 or unc-84 alleles. We will use genome-wide RNAi to screen for more enhancers. The ability to combine genetic, biochemical, and molecular approaches in a developmental system makes C, elegans a powerful system for these studies. Together, these studies will provide mechanistic insight into the fundamental problem of how the nucleus positions itself in the cytoplasm.